Regulatory role of exosomes in colorectal cancer progression and potential as biomarkers

Colorectal cancer(CRC)remains an enormous challenge to human health worldwide.Unfortunately,the mechanism underlying CRC progression is not well understood.Mounting evidence has confirmed that exosomes play a vital role in CRC progression,which has attracted extensive attention among researchers.In addition to acting as messengers between CRC cells,exosomes also participate in the CRC immunomodulatory process and reshape immune function.As stable message carriers and liquid biopsy option under development,exosomes are promising biomarkers in the diagnosis or treatment of CRC.In this review we have described and analyzed the biogenesis and release of exosomes and current research on the role of exosomes in immune regulation and metastasis of CRC.Moreover,we have discussed candidate exosomal molecules as potential biomarkers to diagnose CRC,predict CRC progression,or determine CRC chemoresistance,and described the significance of exosomes in the immunotherapy of CRC.This review provides insight to further understand the role of exosomes in CRC progression and identify valuable biomarkers that facilitate the clinical management of CRC patients.

Effect of Electromagnetic Pulse Exposure on Brain Micro Vascular Permeability in Rats

Objective To observe the effect of electromagnetic pulse (EMP) exposure on cerebral micro vascular permeability in rats. Methods The whole-body of male Sprague-Dawley rats were exposed or sham exposed to 200 pulses or 400 pulses (1 Hz) of EMP at 200 kV/m. At 0.5, 1, 3, 6, and 12 h after EMP exposure, the permeability of cerebral micro vascular was detected by transmission electron microscopy and immunohistochemistry using lanthanum nitrate and endogenous albumin as vascular tracers, respectively. Results The lanthanum nitrate tracer was limited to the micro vascular lumen with no lanthanum nitrate or albumin tracer extravasation in control rat brain. After EMP exposure, the lanthanum nitrate ions reached the tight junction, basal lamina and pericapillary tissue. Similarly, the albumin immunopositive staining was identified in pericapillary tissue. The changes in brain micro vascular permeability were transient, the leakage of micro vascular vessels appeared at 1 h, and reached its peak at 3 h, and nearly recovered at 12 h, after EMP exposure. In addition, the leakage of micro vascular was more obvious after exposure of EMP at 400 pulses than after exposure of EMP at 200 pulses. Conclusion Exposure to 200 and 400 pulses (1 Hz) of EMP at 200 kV/m can increase cerebral micro vascular permeability in rats, which is recoverable.

Effect of Electromagnetic Pulse Exposure on Permeability of Blood-testicle Barrier in Mice

Objective To study the effect of electromagnetic pulse （EMP） exposure on the permeability of blood-testicle barrier （BTB） in mice. Methods Adult male BALB/c mice were exposed to EMP at 200 kV/m for 200 pulses with 2 seconds interval. The mice were injected with 2% Evans Blue solution through caudal vein at different time points after exposure, and the permeability of BTB was monitored using a fluorescence microscope. The testis sample for the transmission electron microscopy was prepared at 2 h after EMP exposure. The permeability of BTB in mice was observed by using Evans Blue tracer and lanthanum nitrate tracer. Results After exposure, cloudy Evans Blue was found in the testicle convoluted seminiferous tubule of mice. Lanthanum nitrate was observed not only between testicle spermatogonia near seminiferous tubule wall and sertoli cells, but also between sertoli cells and primary spermatocyte or secondary spermatocyte. In contrast, lanthanum nitrate in control group was only found in the testicle sertoli cells between seminiferous tubule and near seminifdrous tubule wall. Conclusion EMP exposure could increase the permeability of BTB in the mice.

Damage to Hippocampus of Rats after Being Exposed to Infrasound

Objective The objective was to observe damage of hippocampus in rats after exposure to infrasound, and to assess HSP70 expression in hippocampus. Methods SD rats in the experimental group were exposed to 140 d B（8 Hz） infrasound for 2 h per day for 3 days. The morphology of the hippocampus was examined by transmission electronic microscopic（TEM）. Cell apoptosis was observed by TUNEL staining at 0 h, 24 h, 48 h, and 2 w after exposure. HSP70 expression was detected by immunohistochemistry（IHC） and Western blotting（WB）. Results TEM showed that hippocampus was significantly damaged by exposure, and exhibited recovery 1 week after exposure. The TUNEL data showed that neuronal apoptosis after exposure was significantly higher than in the control rats at 24 h and 48 h, and the apoptotic cells decreased one week after exposure. IHC and WB showed HSP70 expression was significantly higher in the exposed rats, peaked at 24 h. Conclusion Exposure to 140 d B（8 Hz） infrasound for 2 h per day for 3 days appeared to induce damage to the hippocampus of rats, based on changes in ultrastructure and increased cell apoptosis. However, recovery from the damage occurred overtime. HSP70 expression also increased after the exposure and decreased by 48 h.

Altered Expression of Matrix Metalloproteinases and Tight Junction Proteins in Rats following PEMF-Induced BBB Permeability Change

Objective To investigate the expression of occludin,ZO‐1,MMP‐2,and MMP‐9 in cerebral microvasculature following Pulse Electromagnetic Field（PEMF） induced BBB permeability change.Methods Sprague‐Dawley rats were randomized into PEMF and sham exposed groups（n=8）.After exposure to PEMF at 0.5,1,3,6,and 12 h,BBB permeability was measured by Evans‐Blue extravasation.The expression of occludin,ZO‐1,MMP‐2,and MMP‐9 were detected by real‐time quantitative reverse transcriptase PCR and western blotting.MMP‐2 and MMP‐9 activity were detected by EnzChek gelatinase assay.Results Compared with the sham group,PEMF exposure led to increased permeability of the BBB to EB,which was prolonged after exposure.BBB permeability became progressively more severe,and recovered at 6 h.The gene and protein expression of occludin and ZO‐1 were significantly decreased,while MMP‐2 and MMP‐9 expression were significantly increased after exposure to PEMF.All levels of expression recovered 12 h following PEMF.Conclusion Changes to BBB permeability were related to the alteration expression of tight junction proteins and matrix metalloproteinase after exposure to PEMF.

Diabetes mellitus may affect the long-term survival of hepatitis B virus-related hepatocellular carcinoma patients after liver transplantation

AIM to determine whether diabetes mellitus(DM) affects prognosis/recurrence after liver transplantation(Lt) for hepatitis B virus(HBV)-related hepatocellular carcinoma(HCC). METHODS A retrospective study was conducted between January 2000 and August 2013 on 1631 patients with HBV-related HCC who underwent Lt with antiviral prophylaxis. Patient data were obtained from the China Liver transplant Registry(https://www.cltr.org/). to compare the outcomes and tumor recurrence in the HBV-related HCC patients with or without DM, statistical analyses were conducted using χ2 tests, Mann-Whitney tests, the Kaplan-Meier method, log-rank tests and multivariate step-wise Cox regression analysis. RESULTS Univariate analysis of 1631 patients who underwent Lt found overall 1-, 3- and 5-year survival rates of 79%, 73% and 71% respectively in the DM patients, and 84%, 78% and 76% in the non-DM patients respectively. Overall survival rate differences after Lt between the two groups were significant(P = 0.041), but recurrence-free survival rates were not(P = 0.096). By stratified analysis, the overall survival rates in DM patients for age > 50 years(P = 0.002), the presence of vascular invasion(P = 0.096), tumors ≤ 3 cm(P = 0.047), two to three tumor nodules(P = 0.007), Child-Pugh grade B(P = 0.018), and preLt alanine aminotransferase levels between 40 and 80 IU/L(P = 0.017) were significantly lower than in non-DM patients. Additionally, serum α-fetoprotein level > 2000 ng/m L(P = 0.052) was associated with a significant survival difference trend between DM and non-DM patients. Multivariate analysis showed that the presence of DM(P < 0.001, HR = 1.591; 95%CI: 1.239-2.041) was an independent predictor associated with poor survival after Lt. CONCLUSION HBV-related HCC patients with DM have decreased long-term overall survival and poor Lt outcomes. Prevention strategies for HCC patients with DM are recommended.

HMGB1 induces secretion of matrix vesicles which participate in microcalcification of atherosclerotic plaques

AIM: Early calcification of atherosclerotic plaques are colocalized with macrophage and high mobility group box 1( HMGB1),a cytokine associated with biomineralizing process under physiological and pathological conditions. Our study aims to evaluate whether HMGB1 induces ectopic mineralization via promoting the secretion of matrix vesicles( MVs) from macrophages. METHODS: HMGB1 was added to the medium of macrophages,the secretion of MVs in the supernatant was tested by flow cytometry analysis. The mineral deposition in calcifying medium was detected by Alizarin Red staining and von Kossa staining. Transmission electron microscopy showed the formation of hydroxyapatite crystals in MVs. Then we subcutaneous injection into mice with MVs to induce regional mineralization. RESULTS: HMGB1 significantly promoted secretion of MVs from macrophages as raveled by flow cytometry analysis. TNAP activity,considered as a marker of MVs maturation,was higher in HMGB1-induced MVs compared to the control-MVs. HMGB1-MVs also led to mineral deposition in an in vitro MVs-collagen mineralization model. Subcutaneous injection into mice with MVs derived from HMGB1-treated cells showed a greater potential to initiate regional mineralization. Mechanistic experiments revealed that HMGB1 activated neutral sphingomyelinase 2( n SMase2) that involved the receptor for advanced glycation end products( RAGE) and p38MAPK( upstream of n SMase2). Inhibition of n SMase2 with GW4869 or p38 MAPK with SB-239063 prevented MVs secretion and mineral deposition. CONCLUSIONS: HMGB1 induces MVs secretion from macrophages at least in part,via the RAGE / p38 MAPK /n SMase2 signaling pathway. Our findings thus reveal a novel mechanism by which HMGB1 may participated in the early calcification of atherosclerotic plaques.

Risk factors affecting the mother-to-infant transmission of hepatitis B virus: a meta analysis

Objective:To search for risk factors that affect mother-to-infant transmission of hepatitis B virus (HBV). Methods: To obtain studies eligible for meta-analysis, China biological medicine discs and MEDLINE citations were surveyed. Mother HBV DNA or HBeAg positivity, neonate HBeAg positivity, mode of delivery, threatened abortion and threatened premature labor were processed with meta analysis. Criteria for selection of published studies for meta analysis were based on principle by Abdolmaleky HM[1]. Odds ratio (OR) was calculated and summarized by fixed effect model or random-effects model using RevMan software. The heterogeneity of the group of ORs was assessed using an x2 test. The significance of the pooled OR was determined by the u-test. The strength of association was assessed using the OR. An OR>1. 0 indicated a positive association between the risk factor and neonate HBV infection. Results: After meta analysis of factors concerned, a significant association was found between the positivity of HBeAg in mother and neonate, of HBV DNA in mother peripheral serum, and HBV mother-to-infant transmission, with a pooled OR equal to 19.43 (95% CI=8. 77-43. 06), 36.5 (95% 01=19.85-67.11), and 36. 5 (95% CI=19. 85-67. 11) respectively. Mode of delivery, threatened abortion and threatened premature labor proved not to be of risk factors on the mother-to-infant transmission of HBV. Conclusion: Mother HBV DNA or HBeAg positivity and neonate HBeAg positivity were proved to be of risk factors affecting the transmission of HBV from mother to fetal.

Gene Expression Changes in the Pituitary Gland of Rats Exposed to Electromagnetic Pulses

Objective We examined alterations in the expression of tumorigenesis‐related genes in the pituitary gland of rats exposed to electromagnetic pulses （EMP）.Methods The global gene expression profiles of the pituitary gland in EMP‐exposed and control groups were detected by cDNA microarray analysis.We then validated and further investigated the reduced expression of two tumorigenesis‐related genes,Pten,and Jund,by assessing their mRNA and protein expression by quantitative real‐time‐PCR,western blotting,and immunohistochemistry in the pituitary gland of rats 6 months after exposure to EMP.Results EMP exposure induced genome‐wide gene expression changes in the rat pituitary gland.There was decreased expression of the Pten and Jund mRNAs and proteins in EMP‐exposed rats compared with in unexposed control animals.Conclusion EMP exposure alters the expression of tumorigenesis‐related genes in the pituitary gland.These tumorigenesis‐related genes are potentially involved in the development of pituitary gland tumors in rats.

Application of geographical information system technology to epidemiological surveillance and prevention and cure decision-making for SARS

Objective:To expound geographical information system (GIS) technology is a very important tool when it was employed to assist to present the distribution by time and place and the model of transmission of infectious disease. Methods: We illustrated the assistant decision-making support function of GIS with an example of the spatial decision support system for SARS controlling in Shaanxi province of China which was developed by us. Results: The spatial decision support system established by applying GIS technology fulfilled the needs of real-time collection and management and dissemination SARS information and of surveillance and analysis the epidemic situation of SARS. Conclusion: Occurrence and epidemic of diseases, implement prevention and intervention measures and collocation hygienic resources are all with the characteristic of the variation of time and space, therefore, GIS technology has become a powerful tool for identifying risk factors of diseases, providing clues of causation of diseases , evaluating the effects of intervention measures and drawing a health management plan.

The Effects of Electromagnetic Pulse on the Protein Levels of Tight Junction Associated-Proteins in the Cerebral Cortex,Hippocampus,Heart,Lung,and Testis of Rats

Objective To investigate changes in the expression of tight junction （TJ） proteins in the cerebral cortex,hippocampus,heart,lung,and testes of rats after exposure to electromagnetic pulse （EMP）.Methods Eighteen adult male Sprague-Dawley rats were divided into sham and exposure groups.The exposure groups received EMP at 200 kV/m for 200 pulses with a repetition rate of 1 Hz.The expression of TJ proteins （ZO-1,occludin,actin） in the several organs was examined by western blotting.Results ZO-1 levels in the cerebral cortex decreased 1 h and 3 h after EMP exposure compared with sham group （P0.05）.No significant difference was observed for occludin and actin.ZO-1 levels in the hippocampus increased 1 h and 3 h post-exposure （P0.05）,and occludin decreased after 3 h （P0.05）;however,actin was unaffected.ZO-1 levels in the heart increased 3 h post-exposure （P0.05）,occludin decreased 3 h post-exposure （P0.05）,and actin increased 1 h and 3 h post-exposure （P0.05）.ZO-1,occludin and actin levels in the lung decreased compared with those in the sham group （P0.05）.ZO-1 and occludin levels in the testes decreased 1 h and 3 h post-exposure （P0.05）,but actin showed no significant change.Conclusion Exposure to EMP altered the expression levels of TJ proteins,particularly ZO-1,in the organs of adult male rats,which may induce changes in barrier structure and function.

Establishment of management information system for SARS surveillance and control in Shaanxi province of China

Objective: To develop the management information system for SARS surveillance and control in Shaanxi province of China responding to the urgent needs for preventing and curing SARS disease. Methods: Based on geographic information system technology, the management information system for SARS disease in Shaanxi province of China was established using "SuperMap Objects 3.0" GIS development platform and Delphi 7.0.Results: The following functions were implemented in the system: the real-time collection and monitoring, management and analysis, dissemination of SARS disease information, and assistant decision-making support for prevention against SARS disease. Conclusion: The system that integrates epidemiology theories and GIS techniques together can provide a scientific, efficient means for monitoring, prevention of SARS disease in the future.

20-Hydroxyecdysone Improves Neuronal Differentiation of Adult Hippocampal Neural Stem Cells in High Power Microwave Radiation-Exposed Rats

Objective The hippocampus is thought to be a vulnerable target of microwave exposure.The aim of the present study was to investigate whether 20-hydroxyecdysone(20E)acted as a fate regulator of adult rat hippocampal neural stem cells(NSCs).Furthermore,we investigated if 20E attenuated high power microwave(HMP)radiation-induced learning and memory deficits.Methods Sixty male Sprague-Dawley rats were randomly divided into three groups:normal controls,radiation treated,and radiation+20E treated.Rats in the radiation and radiation+20E treatment groups were exposed to HPM radiation from a microwave emission system.The learning and memory abilities of the rats were assessed using the Morris water maze test.Primary adult rat hippocampal NSCs were isolated in vitro and cultured to evaluate their proliferation and differentiation.In addition,hematoxylin&eosin staining,western blotting,and immunofluorescence were used to detect changes in the rat brain and the proliferation and differentiation of the adult rat hippocampal NSCs after HPM radiation exposure.Results The results showed that 20E induced neuronal differentiation of adult hippocampal NSCs from HPM radiation-exposed rats via the Wnt3a/β-catenin signaling pathway in vitro.Furthermore,20E facilitated neurogenesis in the subgranular zone of the rat brain following HPM radiation exposure.Administration of 20E attenuated learning and memory deficits in HPM radiation-exposed rats and frizzled-related protein(FRZB)reduced the 20E-induced nuclear translocation ofβ-catenin,while FRZB treatment also reversed 20E-induced neuronal differentiation of NSCs in vitro.Conclusion These results suggested that 20E was a fate regulator of adult rat hippocampal NSCs,where it played a role in attenuating HPM radiation-induced learning and memory deficits.

西安市老年男性冠心病危险因素队列研究

目的 前瞻性研究老年人群冠心病死亡与常见危险因素的关系 .方法 驻西安市部队干休所男性离休干部 12 6 8名为研究队列 ,自 1987年起对常见冠心病危险因素 (烟酒嗜好、血压、血脂水平、家族史等 )及死亡情况进行 10 a随访 .结果 单因素分析示吸烟、负性生活事件、高血压病史是老年男性冠心病死亡的危险因素 ,相对危险度 (RR)分别为 :2 .2 7(1.6 3～ 3.0 2 ) ,2 .31(1.12～ 2 .5 7) ,5 .5 0 (3.18～ 7.0 4) . Cox风险比例模型分析示吸烟、负性生活事件、年龄入选 ,高血压病史、心血管病史、心血管病家族史、冠心病史、胆固醇和三酰甘油水平未能进入模型 .结论 在老年人群中 ,吸烟、高血压等常见冠心病危险因素亦是冠心病死亡的主要危险因素 .

Breast Cancer Resistance Protein Expression and 5-Fluorouracil Resistance

Objective To filtrate breast cancer resistance protein （BCRP）-mediated resistant agents and to investigate clinical relationship between BCRP expression and drug resistance. Methods MTT assay was performed to filtrate BCRP-mediated resistant agents with BCRP expression cell model and to detect chemosensitivity of breast cancer tissue specimens to these agents. A high performance liquid chromatography （HPLC） assay was established, and was used to measure the relative dose of intracellular retention resistant agents. RT-PCR and immunohistochemistry （IHC） were employed to investigate the BCRP expression in breast cancer tissue specimens. Results MTT assay showed that the expression of BCRP increased with the increasing resistance of 5-fluorouracil （5-Fu） （P〈0.05, n=3） in the cell model, while HPLC assay indicated that the intracellular retention dose of 5-Fu was significantly correlated with the expression of BCRP （t=-0.897, P〈0.05, n=3）. A total of 140 breast cancer tissue specimens were collected. BCRP-positive expression was detected in forty-seven specimens by both RT-PCR and IHC. As shown by MTT assay subsequently, the resistance index （RI） of 47 BCRP-positive breast cancer tissue specimens to 5-Fu was 7-12 times as high as that of adjacent normal tissue samples. BCRP expression was related to 5-Fu resistance （R2=0.8124, P〈0.01）. Conclusion Resistance to 5-Fu can be mediated by BCRR Clinical chemotherapy for breast cancer patients can be optimized based on BCRP-positive expression.

Autophagy Attenuates MnCl2-induced Apoptosis in Human Bronchial Epithelial Cells

Objective To investigate the role of autophagy in MnC l2-induced apoptosis in human bronchial epithelial 16 HBE cells.Methods Cell proliferation was measured by MTT assay.Mitochondrial membrane potential（MMP） and apoptosis were measured by flow cytometry.Autophagic vacuoles were detected by fluorescence microscopy.Cellular levels of apoptosis and autophagy-related proteins were measured by western blotting.Results 16 HBE cell proliferation was inhibited by Mn Cl2 in a dose-and time-dependent manner.Mn Cl2-induced 16 HBE cell growth inhibition was related to MMP depolarization prior to the induction of apoptosis.Our data revealed that Mn Cl2-induced apoptosis in 16 HBE cells was mediated by decreased expression of Bcl-2 and increased levels of cleaved caspase-3.It was observed that when we exposed 16 HBE cells to MnCl2 in a dose-dependent manner,the formation of autophagic vacuoles and the levels of LC-3B-II were elevated.RNA interference of LC3 B in these Mn Cl2-exposed cells demonstrated that MMP loss and apoptosis were enhanced.Additionally,the pan-caspase inhibitor Z-VAD-FMK increased the cellular levels of Bcl-2 and decreased apoptosis,but did not affect the cellular levels of LC3 B in Mn Cl2-treated 16 HBE cells.Conclusion Mn Cl2 dose-and time-dependently inhibits 16 HBE cell proliferation and induces MMP loss and apoptosis.Autophagy acts in a protective role against Mn Cl2-induced apoptosis in 16 HBE cells.

Probability Prediction in Multistate Survival Models for Patients with Chronic Myeloid Leukaemia

In order to find an appropriate model suitable for a multistate survival experiment, 634 patients with chronic myeloid leukaemia (CML) were selected to illustrate the method of analysis. After transplantation, there were 4 possible situations for a patient: disease free, relapse but still alive, death before relapse, and death after relapse. The last 3 events were considered as treatment failure. The results showed that the risk of death before relapse was higher than that of the relapse, especially in the first year after transplantation with competing-risk method. The result of patients with relapse time less than 12 months was much poor by the Kaplan-Meier method. And the multistate survival models were developed, which were detailed and informative based on the analysis of competing risks and Kaplan-Meier analysis. With the multistate survival models, a further analysis on conditional probability was made for patients who were disease free and still alive at month 12 after transplantation. It was concluded that it was possible for an individual patient to predict the 4 possible probabilities at any time. Also the prognoses for relapse either death or not and death either before or after relapse may be given. Furthermore, the conditional probabilities for patients who were disease free and still alive in a given time after transplantation can be predicted.

Lung sounds auscultation technology based on ANC - ICA algorithm in high bat- tlefield noise environment

AIM:To explore the more accurate lung sounds auscultation technology in high battlefield noise environment.METHODS: In this study, we restrain high background noise using a new method-adaptive noise canceling based on independent component analysis (ANC-ICA), the method, by incorporating both second-order and higher-order statistics can remove noise components of the primary input signal based on statistical independence.RESULTS:The algorithm retained the local feature of lung sounds while eliminating high background noise, and performed more effectively than the conventional LMS algorithm.CONCLUSION:This method can cancel high battlefield noise of lung sounds effectively thus can help diagnose lung disease more accurately.

Molecular-Genetic Characterization of Human Rotavirus A Strains Circulating in Moscow, Russia(2009–2014)

Enteric viruses are the most common cause of acute gastroenteritis(AGE) in young children and a significant public health problem globally. Hospital admissions of children under 5 years of age with diarrhea are primarily associated with group A rotavirus(RVA) infection. In this retrospective study, the population structure of viruses linked to AGE etiology in young children hospitalized with AGE in Moscow was evaluated, and molecular characterization of RVA strains was performed.Fecal specimens were collected from children under 5 years old hospitalized with AGE between 2009 and 2014 in Moscow, Russia. Multiplex real-time reverse transcription PCR was used to detect enteric viruses and for G/[P]-genotyping of isolated RVAs. Sequencing of RVA VP7 and VP4 cDNA fragments was used to validate the data obtained by PCRgenotyping. The main causes for hospitalization of children with AGE were RVA(40.1%), followed by noroviruses(11.4%), while adenoviruses, astroviruses, sapoviruses, enteroviruses, and orthoreoviruses were detected in 4.7%, 1.9%,1.4%, 1.2%, and 0.2% of samples tested, respectively. Nosocomial infections, predominantly associated with RVAs and noroviruses, were detected in 24.8% of cases and occurred significantly more frequently in younger infants. The predominant RVA genotype was G4 P[8], detected in 38.7% of RVA-positive cases, whereas genotypes G1 P[8], G9 P[8],G3 P[8], and G2P[4] were found in 11.8%, 6.6%, 4.2%, and 3.3% of cases, respectively. Together, the presence of circulating RVA strains with rare VP7 and VP4 gene variants(G6 and P[9]) highlights the need to conduct continuous epidemiological monitoring of RVA infection.

Spinal CCL2 Promotes Pain Sensitization by Rapid Enhancement of NMDA-Induced Currents Through the ERK-GluN2B Pathway in Mouse Lamina Ⅱ Neurons

Previous studies have shown that CCL2(C-C motif chemokine ligand 2)induces chronic pain,but the exact mechanisms are still unknown.Here,we established models to explore the potential mechanisms.Behavioral experiments revealed that an antagonist of extracellular signal-regulated kinase(ERK)inhibited not only CCL2-induced inflammatory pain,but also pain responses induced by complete Freund’s adjuvant.We posed the question of the intracellular signaling cascade involved.Subsequent experiments showed that CCL2 up-regulated the expression of phosphorylated ERK(pERK)and N-methyl D-aspartate receptor[NMDAR]subtype 2B(GluN2B);meanwhile,antagonists of CCR2 and ERK effectively reversed these phenomena.Whole-cell patchclamp recordings revealed that CCL2 enhanced the NMDAR-induced currents via activating the pERK pathway,which was blocked by antagonists of GluN2B and ERK.In summary,we demonstrate that CCL2 directly interacts with CCR2 to enhance NMDAR-induced currents,eventually leading to inflammatory pain mainly through the CCL2-CCR2-pERK-GluN2B pathway.